Dominant negative down-regulation of endotoxin-induced tumor necrosis factor α production byLpsd/Ran

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Abstract
We recently showed that adenoviral transfer and expression of the Lpsd/Ran gene isolated from endotoxin-resistant C3H/HeJ mice could protect endotoxin-sensitive mice from endotoxic shock. Elevation of proinflammatory cytokines, such as tumor necrosis factor α (TNF-α), is thought to be essential for the development of septic shock. To investigate the extent to which Lpsd/Ran affects TNF-α production, we transduced primary macrophages from endotoxin-sensitive and -resistant mice with adenoviral vectors expressing the wild-type and the mutant Lps/Ran cDNAs and other control genes, and compared the amount of TNF-α produced by these various transduced macrophages. Successful transfer and expression of Lpsd/Ran cDNA in endotoxin-sensitive C3H/HeOuJ macrophages reduced TNF-α production upon lipopolysaccharide (LPS) stimulation, as compared with macrophages transduced with vectors expressing the wild-type Lpsn/Ran cDNA, the green fluorescent protein gene, or the lacZ gene. On the other hand, successful transfer and expression of the wild-type Lpsn/Ran cDNA in primary macrophages from endotoxin-resistant C3H/HeJ mice failed to induce TNF-α production to any significant extent unless a very high LPS concentration was used. Given our previous demonstration that Lpsn/Ran functions effectively in restoring LPS responsiveness in B cells from C3H/HeJ mice, we conclude that Lps/Ran is involved in a CD14-independent signal transduction pathway. This dominant negative down-regulation by Lpsd/Ran on TNF-α production by macrophages and probably other innate immune responses may be key to the development of an effective gene therapy for endotoxic or septic shock.